To clarify the role of protein phosphorylation in the regulation of insulin secretion we studied the effects of protein phosphatase inhibitors on insulin selease, glucose metabolism, cytosolic calcium concentration, and cyclic AMP accumulation in rat pancreatic islets. In the batch incubation of intact islets insulin release induced by 10mM glucose was inhibited dose-dependently in the order of calyculin A,okadaic acid, and cantharidine.Additon of diazoxide and high K doubled glucose-induced insuliln release. This release was also inhibited by the three protein phosphatase inhibitors with similar values of IC_<50>. Okadaic acid (1muM) did not affect CO_2 production from glucose. Neither calyculin A (0.2muM) nor okadaic acid influenced changes in cytosolic calcium concentration induced by glucose or high K.Okadaic acid inhibied cyclic AMP accumulation by 20% in the presence of 3 or 10mM glucose. In islets permeabilized by beta-escin, an increase in calcium concentration from 10^<-7> to 10^<-5> M increased insulin release by 50%.Okadaic acid increased insulin release by 350% at 10^<-7> M Ca and by 100% at 10^<-5> M Ca. At 10^<-7> M Ca forskolin (1muM) did not induce insulin release and further addition of okadaic acid increased it to a similar level to okadaic acid alone. In contrast TPA (162nM) increased insulin release by 320% and further addtion of okadaic acid augmented it. These data suggest that protein phosphatase 1 may play a role in cyclic AMP accumulation and/or an increase in sensitivity of the secretory machinery to calcium.